- Email: [email protected]
633 Specific activation of Akt3 in ovarian cancer
s190
Tuesday
23 September
2003
Poster Session
The aim of this study was to examine the effect of the ACE inhibitor ramipril (RAM) on tumour growth and metastases as well as the underlying mechanisms. Material and methods: A murine Lewis Lung tumour and metastasismodel was used. C57BU6J mice were inoculated subcutaneously with 2~10~ tumour cells on day 1. Treatment was initiated the following day as RAM 30mgkglday in the drinking water, cyclophosphamide (CTX) IOOmgIkg intraperitoneally (i.p.) day 2, doxorubicin (DOX) 2.4mgikg i.p. day 2-5 or combinations of RAM+CTX or RAM+DOX. Tumour size was recorded three times per week and mice were sacrificed on day 28. The lungs were processed for stereological determination of metastasis volume. Turnouts were collected and examined for expression of MMPs at mRNA and protein levels by RT-PCR and Western blot. Some of the lungs were examined for MMP expression at the protein level as well. Resutts: Based on time to reach a tumour volume of 800mm3 KaplanMeier plots were constructed for each treatment group. Compared to saline-treated controls, RAM treatment significantly increased time to tumour volume 800mm3 (23 davs versus 21 davs. o
631
POSTER
Nitric oxide up- regulates cyciooxygenase-2 expression through the CAMP- response element in its promoter in a head and neck cancer ceil line S.W. Park’, M.W. Sung*. ‘Seoul National University College of Medicine, Cancer Research Institute, Seoul, Korea; ‘Seoul National University College of Medicine, Department of Otorhinolaryngology - Head and Neck Surgem Clinical Research Ins, Seoul, Korea Background: We previously observed the over-expression of cyclooxygenase-2 (COX-2) and the increased production of prostaglandin E by nitric oxide (NO) in several cancer cell lines. In this study, we investigated the mechanisms of interaction between the NO and COX-2 pathways in head and neck cancer cells. Material and Methods: For our experiments, we used plasmids containing partial COX-2 promoter region and the fusion franeactivator plasmids (pFA-ATF-2, pFA2-CREB, and pFA2-cJun from stratagene) and performed western blotting and promoter-luciferase assay. Results: CAMP-response element (CRE) was identified as a critical factor of COX-2 expression in SNU-1041. It was found that at least three transcription factors (TFs) - CREB, ATF-2 and c-jun, could bind to CRE of the COX-2 promoter and that their activities were increased by SNAP, a NO donor. Also we found that the activation of soluble guanylate cyclase, p38 and JNK by NO might play an important role in COX-2 over-expression throuah the up-reaulation of these three TFs. The effect of dibutvrvl-cGMP on COX-2 expre&ion was similar to that of SNAP and was blockebby a p38 inhibitor, not by a JNK inhibitor. In addition, we found that dibutyryl-cGMP might activate CREB and ATF-2, whose activities were increased by ~38, but not c-jun. Moreover, NO induced JNK signaling followed by the activation of c-jun and ATF-2 in cGMP-independent manner. Conclusions: These results imply that NO generated endogenously at low concentrations may affect many gene expressions, including COX-2, which can promote the growth and survival of tumor cells.
632
POSTER
Genomic DNA amplification of Decoy receptor 3 (DcR3) correlates with cancer progression of well-differentiated coiorectal adenocarcinoma. M. Kawahara, Y. Koyama, C. Kanbayashi, V. Valera, T. liai, l-f. Okamoto, T. Suda, K. Hatakeyama. Niigafa University Graduate School of Medical and, Division of Digestive and Genera/ Surgery, Niigata, Japan Background: Decoy receptor 3 (DcR3), a member of tumor necrosis factor receptor (TNFR) superfamily, shows inhibitory effect on Fas- mediated apoptosis. We have reported the relationship between DcR3 mRNA overexpression and the progression of well-differentiated colorectal adenocarcinoma (17th Meeting of the European Association for Cancer Research, abstract #403). In the present study, we examined the relationship between DcR3 gene amplification and Fas mRNA expression, and also examined the correlation of DcR3 gene amplification with apoptotic cell death to clarify its effect(s) in human well- differentiated colorectal adenocarcinoma. Materials and methods: Tissue specimens were obtained from 27 patients with well-differentiated colorectal adenocarcinoma who underwent operations at Niigata University Hospital between 1998 and 2002. Genomic DNA of cancer tissue was extracted from paraffin embedded sections by microdissection under light microscope. Total cellular RNA, extracted from tissue samples of both cancer and nomal mucosa, were reversetranscribed to synthesize cDNA. Quantitative real- time PCR was carried out to determine genomic DNA amplification of DcR3 and mRNA expression of Fas by standardizing with b-globin gene. In 19 patients, cancer cell death was examined visibly by in situ enzymatic labeling of DNA strand breaks using Apoptag in situ oligo ligation (ISOL) kit. Statistical analysis was performed by Mann-Whitney U-test, Kruskal- Wallis test, and Spearman’s correlation coefficient by the rank test, and the statistical significance was defined as PxO.05. Results: Genomic DNA amplification of DcR3 was found in 23 cases (85.2%), and was significantly increased in patients with tumor invasion deeper than subserosa or non-peritonealized perirectal tissues (P=O.O014), and in the vascular invasion-positive patients (P=O.O14). There was no significant correlation between DcR3 gene amplification and other clinicopathological features including Fas mRNA expression. By ISOL in situ cell death detection assay, DcR3 gene amplification was significantly increased in apoptotic cells-negative patients compared to positive patients (P=O.O412). Conclusions: These results suggest that DcR3 gene amplification in well-differentiated colorectal adenocarcinoma may be one of the factors for evasion of apoptosis, and may be involved in cancer progression effecting variables such as depth of tumor invasion and vascular invasion.
633
Specific activation
POSTER
of Akt3 in ovarian cancer
B.E. Cristiano, N. Marmy Conus, K.M. Hannan, I. Campbell, R.B. Pearson. Peter MacCallum Cancer Cenrre, Trescowthick Research Laboratories, East Melbourne, Australia Background: The serine/threonine protein kinase Akt exists as three isoforms; Aktl, AM2 and Akt3. Akt, which is activated in response to mitogenic stimuli, may contribute to tumourigenesis at multiple levels with the kinase shown to play prominent roles in several processes considered hallmarks of cancer including the regulation of proliferation, cell survival, invasiveness and angiogenesis. Recent studies suggest deregulation of specific Akt isoforms may be involved in individual tumour types including ovarian, breast and pancreatic cancers. Levels of Aktl and Akt2 activity have been shown to be amplified in 6 and 36% of primary ovarian tumours, respectively. However, this work has been limited by lack of specific reagents for the Akt isoforms. This study has investigated the role of Akt3, in parallel with Aktl and Akt2, in ovarian cancer. Materials and Methods: The expression of each AM isoform was assessed by western blotting using isoform specific antibodies. AM activity was determined by direct assay using the specific peptide substrate RPRAATF, by isoform-specific immunoprecipitation assays and by western blotting with phosphospecific antibodies. Expression of Akt2, Akt3 and activated Akt (representing all 3 isoforms) was assessed in primary ovarian tumour samples by immunohistochemistry. Results: A screen of isoform expression in 8 ovarian cancer cell lines and a non-tumourigenic control cell line, revealed Aktl expression to vary across the cell lines, whereas Akt2 was detected in only one cell line (OVCARS). Expression of Akt3 also varied across the cell lines with marked overexpression in 2 of 9 cell lines tested (OVCA429 and DOV13). Total Akt and Akt3 specific activity was shown to correlate with overexpression of
Paediatric
Tuesday
oncology
Akt3 in these cell lines. This increased activity also correlated with faster proliferation rates and increased survival in the absence of serum. Treatment of the cells with the Pl3K inhibitor LY294002 resulted in increased cell death in those cells with high Akt activity. Akt isoform expression was assessed in 53 primary ovarian tumour samples revealing high Akt3 expression in 33% mutinous, 59% serous, and 66.6% endometrioid tumours. High Akt2 expression was observed in 11% mutinous, 18% serous but not in endometrioid tumours. Positive phospho-S473 staining, representing active AM, correlated with high Akt3 expression. Conclusion: These results suggest that Akt3 may play an important role in ovarian tumourigenesis.
634
The cytochrome carcinogenesis.
POSTER
~450 (CYP) family 1 at early stages of
A. Vaiman. institute of Carcinogenesis, N.B/okhin Cancer Rese, Laboratory of Tumor Cell Genetics, Moscow, Russian Federation
Paediatrie oncology POSTER
Second tumors after treatment for Hodgkin’s in children S. Safonova, A. Malinin, Yu. Punanov. Cancer Paediatric, St. Petersburg, Russian Federation
Research
2003
lymphoma
(HL)
s191
which were include chemotherapy (MOPP, COPP, ABVD, OPPA, PCVP) and radiotherapy (20-50 Gy). Results: Second tumors were found in 20 pts (4,4%). Malignant tumors were in 12 pts (2.7%), and benignant tumors 8 pts (1,8%). Among SMT were: stomack cancer -3, breast cancer -2, thyroid cancer -1, liposarcoma -1, glyosarcoma -1, malignant schwannoma -1, rhabdomyosarcoma -1, acute myeloblastic leukemia -1. SMT were reveal in period from 30 to 340 months to beginning the treatment HL (med. =165 months). Mortality rate for patients with SMT compose 41,7%. Second benignant tumors (SBT) were diagnosed in 8 pts during 46-300 months (med.=153). Thyroid adenoma were in 5 pts, breast fibroadenoma 2, papillomatosis of larynx 1. One patient suffered from 2 SBT thyroid adenoma and neurinoma. Conclusion: SMT more often reveal in patients with HL, which treated in age older 10 years (8 pts from 12) and in women (8 pts from 12). All the second solid tumors (SMT and SBT) are localized in zone of radiotherapy with dose 40 Gy. Thyroid and breast tumors are the most frequent in structure of second tumors.
636
Background: CYPl plays an important role in activation of environmental carcinogens including polycyclic aromatic hydrocarbons, PAH and some drugs. It was suggested that PAH is present in tobacco smoke and participates in tumor development in smokers. There are many studies of CYPs in cancers, however the data on regulation of CYP isoforms at early stages of carcinogenesis are limited. Using two models of cell immortalization and transformation, we studied mRNA expression and activity of CYPI enzymes as well as the expression of AHR and ARNT genes that encode the regulators of CYPl expression. Material and methods: cultivation of cell culture, RNA isolation, RT-PCR, benzo/a/pyrene-hydroxilase activity assay, cell transfection, cell survival assay. Result: In the embryo rat fibroblasts (RF) constitutive level of CYP IAl mRNA was not detectable, whereas CYPlSl mRNA was expressed. After cells immortalization with Rauscher virus (F-27/RLV), mRNA level of CYP 1Ai became high, and CYP 161 level increased in comparison with RF cells. The F-27/RLV cells oxidized benzo/afpyrene more effectively and were more sensitive to toxic effects of benzo/a@yrene and 7.12dimethylbenz/a/anthracene than RF cells. In spontaneously immortalized embryonic rat fibroblasts (Ratl) we found high expression of CYPl67 mRNA compared to RF cells. Treatment with TCDD increased CYP I61 mRNA level in both rat cell lines. Unlike RF, Rauscher immortalized cells with relatively high level of CYPl expression were sensitive to transforming effect of benzo/a/pyrene. In transformed clones levels of CYPlAi and CYPlSl mRNA were lower compared with F-27/RLV cells. Benzo/a/pyrene - hydroxylase activity decreased in transformed cells. In Rat/ras transformed cells obtained after transfection of N-ra~prz gene, the constitutive expression of CYPlSl mRNA disappeared in comparison with Rat1 cells. The mRNAs of proteins which take part in the regulation of enzymatic induction of CYPI (AHR and ARNT) were the same in all cell models studied. Conclusion: Constitutive and inducible levels of CYPfSl mRNA increase after immortalization. Transformation of immortalized cells provokes disappearance of CYPlSl expression. Since AHR and ARNTexpression are similar in all cells studied, we suggest that other factors besides AHR and ARNT take part in CYPl regulation.
635
23 September
POSTER
Neuroblastoma in adolescents and adults: analysis of a mono-institutional series of 33 consecutive patients. M. Podda’, D. Polastri’, M. Massimino’, P. Collini2, L. Gandola3, L. Piva4, M. Terenziani’ , M. Casanova’, F. Fossati Bellani’ , R. Luksch’ ’ lstitufo Nzionale dei Tumori, Pediatric Oncologyr Milano, /ta/y; 2/stituto Nzionale dei Tumori, Pathology, Milano, Italy; 3 lsfituto Nzionale dei Tumori, Radiotherapy, Milano, Ha/y; 4 lstituto Nzionale dei Tumori, Surgery Milano, ha/y Background: Neuroblastoma (NBL) is the most common extra-cranial solid tumor in children. More than 95% of patients at diagnosis are younger than 10 years. Adolescents and adults have a grave prognosis, but may have a more indolent course. Material and methods: From 1980 to 2002, 33 patients (stage I= 3, stage II= 6, stage lll=8, stage IV=16) with newly diagnosed NBL older than 12 have been admitted at lstituto Nazionale dei Tumori of Milan. Median age was 17 yrs (range 12-69); M/F ratio was 1.2. Symptoms were present and disregarded for many months in the majority of cases: the mean time frame between the onset of symptoms and diagnosis was 15 months. Site of the primary tumor was retroperitoneum in 19 cases, mediastinum in 5, pelvis in 1, cervical in 1, while 6 had an esthesioneuroblastoma; 1 case the primary tumor was unknown. LDH was elevated in 15/33 pts. Treatment applied: surgery alone for stage I; post-operative radiotherapy for stage II; stage Ill and IV received chemotherapy regimens including anthracycline + cyclophosphamide + vincristine f ifosfamide f etoposideh cisplatinum. In addition, lo/16 stage IV pts were submitted to sequential hemi-body irradiation as consolidation treatment. Radiotherapy and/or surgery on primary and metastases were decided on individual basis. Results: 20/33 relapsed: 0 stage I, 1 stage II, 4 stage Ill, 13 stage IV, and 18 relapsed died. The median follow-up is 42 months (range 12-264). EFS and OS probability at 5 years are shown in the table:
EFS OS
stage I
stage II
stage III
1 1
0.67 0.83
0.40 0.56
sage
IV
0 0.12
In this series a consistent number of late relapse/progression were observed: time to progression/relapse ranged from 3 to 58 months and the time from relapse to death from 2 to 75 months. In univariate analysis, together with the stage, the only statistically significant prognostic factor is LDH level at diagnosis: an elevated LDH negatively predicted the outcome (5 yrs OS: normal 54%, pathological 0; p 0.0089). Conclusions: Localized NBL (stage I and II) in adolescents and adults have the same good prognosis of children. For pts with locally advanced and metastatic disease, late events were frequently observed, thus suggesting a lower biological aggressiveness of the disease in this subset. Nevertheless, the prognosis for these patients is dismal.
Insfitute,
Second malignant tumors (SMT) are one of the most severe complications in children treated for HL. The aim of the study was to investigate the incidence of second tumors in children with HL. Materials and methods: We observed 450 pts. in CR after treatment of HL from 1972 to 1997. The period after treatment was 60-360 months (med.=134,5). The age of pts was from 2 to 18 years (med.=12,5 yrs; M/F=2,6/1). Histological subtypes nodular sclerosing and mixed cellularity were predominance. The Ill and IV stages of HL were reveal in 70% patients. Practically all children were undergo combined treatment HL,
637
POSTER
Evidence for a redox mechanism of action of prednisolone childhood acute lymphoblastic leukaemia through the identification of the novel gene CGI-31. A. Burke, P. Kearns, N. Goulden. University Microbiolog): Bristol, United Kingdom Glucocorticoids are the most acute lymphoblastic leukaemia
of Bristol,
Pafhology
in
and
important drugs used in the treatment of and poor response to these drugs during
Tuesday
23 September
2003
Poster Session
The aim of this study was to examine the effect of the ACE inhibitor ramipril (RAM) on tumour growth and metastases as well as the underlying mechanisms. Material and methods: A murine Lewis Lung tumour and metastasismodel was used. C57BU6J mice were inoculated subcutaneously with 2~10~ tumour cells on day 1. Treatment was initiated the following day as RAM 30mgkglday in the drinking water, cyclophosphamide (CTX) IOOmgIkg intraperitoneally (i.p.) day 2, doxorubicin (DOX) 2.4mgikg i.p. day 2-5 or combinations of RAM+CTX or RAM+DOX. Tumour size was recorded three times per week and mice were sacrificed on day 28. The lungs were processed for stereological determination of metastasis volume. Turnouts were collected and examined for expression of MMPs at mRNA and protein levels by RT-PCR and Western blot. Some of the lungs were examined for MMP expression at the protein level as well. Resutts: Based on time to reach a tumour volume of 800mm3 KaplanMeier plots were constructed for each treatment group. Compared to saline-treated controls, RAM treatment significantly increased time to tumour volume 800mm3 (23 davs versus 21 davs. o
631
POSTER
Nitric oxide up- regulates cyciooxygenase-2 expression through the CAMP- response element in its promoter in a head and neck cancer ceil line S.W. Park’, M.W. Sung*. ‘Seoul National University College of Medicine, Cancer Research Institute, Seoul, Korea; ‘Seoul National University College of Medicine, Department of Otorhinolaryngology - Head and Neck Surgem Clinical Research Ins, Seoul, Korea Background: We previously observed the over-expression of cyclooxygenase-2 (COX-2) and the increased production of prostaglandin E by nitric oxide (NO) in several cancer cell lines. In this study, we investigated the mechanisms of interaction between the NO and COX-2 pathways in head and neck cancer cells. Material and Methods: For our experiments, we used plasmids containing partial COX-2 promoter region and the fusion franeactivator plasmids (pFA-ATF-2, pFA2-CREB, and pFA2-cJun from stratagene) and performed western blotting and promoter-luciferase assay. Results: CAMP-response element (CRE) was identified as a critical factor of COX-2 expression in SNU-1041. It was found that at least three transcription factors (TFs) - CREB, ATF-2 and c-jun, could bind to CRE of the COX-2 promoter and that their activities were increased by SNAP, a NO donor. Also we found that the activation of soluble guanylate cyclase, p38 and JNK by NO might play an important role in COX-2 over-expression throuah the up-reaulation of these three TFs. The effect of dibutvrvl-cGMP on COX-2 expre&ion was similar to that of SNAP and was blockebby a p38 inhibitor, not by a JNK inhibitor. In addition, we found that dibutyryl-cGMP might activate CREB and ATF-2, whose activities were increased by ~38, but not c-jun. Moreover, NO induced JNK signaling followed by the activation of c-jun and ATF-2 in cGMP-independent manner. Conclusions: These results imply that NO generated endogenously at low concentrations may affect many gene expressions, including COX-2, which can promote the growth and survival of tumor cells.
632
POSTER
Genomic DNA amplification of Decoy receptor 3 (DcR3) correlates with cancer progression of well-differentiated coiorectal adenocarcinoma. M. Kawahara, Y. Koyama, C. Kanbayashi, V. Valera, T. liai, l-f. Okamoto, T. Suda, K. Hatakeyama. Niigafa University Graduate School of Medical and, Division of Digestive and Genera/ Surgery, Niigata, Japan Background: Decoy receptor 3 (DcR3), a member of tumor necrosis factor receptor (TNFR) superfamily, shows inhibitory effect on Fas- mediated apoptosis. We have reported the relationship between DcR3 mRNA overexpression and the progression of well-differentiated colorectal adenocarcinoma (17th Meeting of the European Association for Cancer Research, abstract #403). In the present study, we examined the relationship between DcR3 gene amplification and Fas mRNA expression, and also examined the correlation of DcR3 gene amplification with apoptotic cell death to clarify its effect(s) in human well- differentiated colorectal adenocarcinoma. Materials and methods: Tissue specimens were obtained from 27 patients with well-differentiated colorectal adenocarcinoma who underwent operations at Niigata University Hospital between 1998 and 2002. Genomic DNA of cancer tissue was extracted from paraffin embedded sections by microdissection under light microscope. Total cellular RNA, extracted from tissue samples of both cancer and nomal mucosa, were reversetranscribed to synthesize cDNA. Quantitative real- time PCR was carried out to determine genomic DNA amplification of DcR3 and mRNA expression of Fas by standardizing with b-globin gene. In 19 patients, cancer cell death was examined visibly by in situ enzymatic labeling of DNA strand breaks using Apoptag in situ oligo ligation (ISOL) kit. Statistical analysis was performed by Mann-Whitney U-test, Kruskal- Wallis test, and Spearman’s correlation coefficient by the rank test, and the statistical significance was defined as PxO.05. Results: Genomic DNA amplification of DcR3 was found in 23 cases (85.2%), and was significantly increased in patients with tumor invasion deeper than subserosa or non-peritonealized perirectal tissues (P=O.O014), and in the vascular invasion-positive patients (P=O.O14). There was no significant correlation between DcR3 gene amplification and other clinicopathological features including Fas mRNA expression. By ISOL in situ cell death detection assay, DcR3 gene amplification was significantly increased in apoptotic cells-negative patients compared to positive patients (P=O.O412). Conclusions: These results suggest that DcR3 gene amplification in well-differentiated colorectal adenocarcinoma may be one of the factors for evasion of apoptosis, and may be involved in cancer progression effecting variables such as depth of tumor invasion and vascular invasion.
633
Specific activation
POSTER
of Akt3 in ovarian cancer
B.E. Cristiano, N. Marmy Conus, K.M. Hannan, I. Campbell, R.B. Pearson. Peter MacCallum Cancer Cenrre, Trescowthick Research Laboratories, East Melbourne, Australia Background: The serine/threonine protein kinase Akt exists as three isoforms; Aktl, AM2 and Akt3. Akt, which is activated in response to mitogenic stimuli, may contribute to tumourigenesis at multiple levels with the kinase shown to play prominent roles in several processes considered hallmarks of cancer including the regulation of proliferation, cell survival, invasiveness and angiogenesis. Recent studies suggest deregulation of specific Akt isoforms may be involved in individual tumour types including ovarian, breast and pancreatic cancers. Levels of Aktl and Akt2 activity have been shown to be amplified in 6 and 36% of primary ovarian tumours, respectively. However, this work has been limited by lack of specific reagents for the Akt isoforms. This study has investigated the role of Akt3, in parallel with Aktl and Akt2, in ovarian cancer. Materials and Methods: The expression of each AM isoform was assessed by western blotting using isoform specific antibodies. AM activity was determined by direct assay using the specific peptide substrate RPRAATF, by isoform-specific immunoprecipitation assays and by western blotting with phosphospecific antibodies. Expression of Akt2, Akt3 and activated Akt (representing all 3 isoforms) was assessed in primary ovarian tumour samples by immunohistochemistry. Results: A screen of isoform expression in 8 ovarian cancer cell lines and a non-tumourigenic control cell line, revealed Aktl expression to vary across the cell lines, whereas Akt2 was detected in only one cell line (OVCARS). Expression of Akt3 also varied across the cell lines with marked overexpression in 2 of 9 cell lines tested (OVCA429 and DOV13). Total Akt and Akt3 specific activity was shown to correlate with overexpression of
Paediatric
Tuesday
oncology
Akt3 in these cell lines. This increased activity also correlated with faster proliferation rates and increased survival in the absence of serum. Treatment of the cells with the Pl3K inhibitor LY294002 resulted in increased cell death in those cells with high Akt activity. Akt isoform expression was assessed in 53 primary ovarian tumour samples revealing high Akt3 expression in 33% mutinous, 59% serous, and 66.6% endometrioid tumours. High Akt2 expression was observed in 11% mutinous, 18% serous but not in endometrioid tumours. Positive phospho-S473 staining, representing active AM, correlated with high Akt3 expression. Conclusion: These results suggest that Akt3 may play an important role in ovarian tumourigenesis.
634
The cytochrome carcinogenesis.
POSTER
~450 (CYP) family 1 at early stages of
A. Vaiman. institute of Carcinogenesis, N.B/okhin Cancer Rese, Laboratory of Tumor Cell Genetics, Moscow, Russian Federation
Paediatrie oncology POSTER
Second tumors after treatment for Hodgkin’s in children S. Safonova, A. Malinin, Yu. Punanov. Cancer Paediatric, St. Petersburg, Russian Federation
Research
2003
lymphoma
(HL)
s191
which were include chemotherapy (MOPP, COPP, ABVD, OPPA, PCVP) and radiotherapy (20-50 Gy). Results: Second tumors were found in 20 pts (4,4%). Malignant tumors were in 12 pts (2.7%), and benignant tumors 8 pts (1,8%). Among SMT were: stomack cancer -3, breast cancer -2, thyroid cancer -1, liposarcoma -1, glyosarcoma -1, malignant schwannoma -1, rhabdomyosarcoma -1, acute myeloblastic leukemia -1. SMT were reveal in period from 30 to 340 months to beginning the treatment HL (med. =165 months). Mortality rate for patients with SMT compose 41,7%. Second benignant tumors (SBT) were diagnosed in 8 pts during 46-300 months (med.=153). Thyroid adenoma were in 5 pts, breast fibroadenoma 2, papillomatosis of larynx 1. One patient suffered from 2 SBT thyroid adenoma and neurinoma. Conclusion: SMT more often reveal in patients with HL, which treated in age older 10 years (8 pts from 12) and in women (8 pts from 12). All the second solid tumors (SMT and SBT) are localized in zone of radiotherapy with dose 40 Gy. Thyroid and breast tumors are the most frequent in structure of second tumors.
636
Background: CYPl plays an important role in activation of environmental carcinogens including polycyclic aromatic hydrocarbons, PAH and some drugs. It was suggested that PAH is present in tobacco smoke and participates in tumor development in smokers. There are many studies of CYPs in cancers, however the data on regulation of CYP isoforms at early stages of carcinogenesis are limited. Using two models of cell immortalization and transformation, we studied mRNA expression and activity of CYPI enzymes as well as the expression of AHR and ARNT genes that encode the regulators of CYPl expression. Material and methods: cultivation of cell culture, RNA isolation, RT-PCR, benzo/a/pyrene-hydroxilase activity assay, cell transfection, cell survival assay. Result: In the embryo rat fibroblasts (RF) constitutive level of CYP IAl mRNA was not detectable, whereas CYPlSl mRNA was expressed. After cells immortalization with Rauscher virus (F-27/RLV), mRNA level of CYP 1Ai became high, and CYP 161 level increased in comparison with RF cells. The F-27/RLV cells oxidized benzo/afpyrene more effectively and were more sensitive to toxic effects of benzo/a@yrene and 7.12dimethylbenz/a/anthracene than RF cells. In spontaneously immortalized embryonic rat fibroblasts (Ratl) we found high expression of CYPl67 mRNA compared to RF cells. Treatment with TCDD increased CYP I61 mRNA level in both rat cell lines. Unlike RF, Rauscher immortalized cells with relatively high level of CYPl expression were sensitive to transforming effect of benzo/a/pyrene. In transformed clones levels of CYPlAi and CYPlSl mRNA were lower compared with F-27/RLV cells. Benzo/a/pyrene - hydroxylase activity decreased in transformed cells. In Rat/ras transformed cells obtained after transfection of N-ra~prz gene, the constitutive expression of CYPlSl mRNA disappeared in comparison with Rat1 cells. The mRNAs of proteins which take part in the regulation of enzymatic induction of CYPI (AHR and ARNT) were the same in all cell models studied. Conclusion: Constitutive and inducible levels of CYPfSl mRNA increase after immortalization. Transformation of immortalized cells provokes disappearance of CYPlSl expression. Since AHR and ARNTexpression are similar in all cells studied, we suggest that other factors besides AHR and ARNT take part in CYPl regulation.
635
23 September
POSTER
Neuroblastoma in adolescents and adults: analysis of a mono-institutional series of 33 consecutive patients. M. Podda’, D. Polastri’, M. Massimino’, P. Collini2, L. Gandola3, L. Piva4, M. Terenziani’ , M. Casanova’, F. Fossati Bellani’ , R. Luksch’ ’ lstitufo Nzionale dei Tumori, Pediatric Oncologyr Milano, /ta/y; 2/stituto Nzionale dei Tumori, Pathology, Milano, Italy; 3 lsfituto Nzionale dei Tumori, Radiotherapy, Milano, Ha/y; 4 lstituto Nzionale dei Tumori, Surgery Milano, ha/y Background: Neuroblastoma (NBL) is the most common extra-cranial solid tumor in children. More than 95% of patients at diagnosis are younger than 10 years. Adolescents and adults have a grave prognosis, but may have a more indolent course. Material and methods: From 1980 to 2002, 33 patients (stage I= 3, stage II= 6, stage lll=8, stage IV=16) with newly diagnosed NBL older than 12 have been admitted at lstituto Nazionale dei Tumori of Milan. Median age was 17 yrs (range 12-69); M/F ratio was 1.2. Symptoms were present and disregarded for many months in the majority of cases: the mean time frame between the onset of symptoms and diagnosis was 15 months. Site of the primary tumor was retroperitoneum in 19 cases, mediastinum in 5, pelvis in 1, cervical in 1, while 6 had an esthesioneuroblastoma; 1 case the primary tumor was unknown. LDH was elevated in 15/33 pts. Treatment applied: surgery alone for stage I; post-operative radiotherapy for stage II; stage Ill and IV received chemotherapy regimens including anthracycline + cyclophosphamide + vincristine f ifosfamide f etoposideh cisplatinum. In addition, lo/16 stage IV pts were submitted to sequential hemi-body irradiation as consolidation treatment. Radiotherapy and/or surgery on primary and metastases were decided on individual basis. Results: 20/33 relapsed: 0 stage I, 1 stage II, 4 stage Ill, 13 stage IV, and 18 relapsed died. The median follow-up is 42 months (range 12-264). EFS and OS probability at 5 years are shown in the table:
EFS OS
stage I
stage II
stage III
1 1
0.67 0.83
0.40 0.56
sage
IV
0 0.12
In this series a consistent number of late relapse/progression were observed: time to progression/relapse ranged from 3 to 58 months and the time from relapse to death from 2 to 75 months. In univariate analysis, together with the stage, the only statistically significant prognostic factor is LDH level at diagnosis: an elevated LDH negatively predicted the outcome (5 yrs OS: normal 54%, pathological 0; p 0.0089). Conclusions: Localized NBL (stage I and II) in adolescents and adults have the same good prognosis of children. For pts with locally advanced and metastatic disease, late events were frequently observed, thus suggesting a lower biological aggressiveness of the disease in this subset. Nevertheless, the prognosis for these patients is dismal.
Insfitute,
Second malignant tumors (SMT) are one of the most severe complications in children treated for HL. The aim of the study was to investigate the incidence of second tumors in children with HL. Materials and methods: We observed 450 pts. in CR after treatment of HL from 1972 to 1997. The period after treatment was 60-360 months (med.=134,5). The age of pts was from 2 to 18 years (med.=12,5 yrs; M/F=2,6/1). Histological subtypes nodular sclerosing and mixed cellularity were predominance. The Ill and IV stages of HL were reveal in 70% patients. Practically all children were undergo combined treatment HL,
637
POSTER
Evidence for a redox mechanism of action of prednisolone childhood acute lymphoblastic leukaemia through the identification of the novel gene CGI-31. A. Burke, P. Kearns, N. Goulden. University Microbiolog): Bristol, United Kingdom Glucocorticoids are the most acute lymphoblastic leukaemia
of Bristol,
Pafhology
in
and
important drugs used in the treatment of and poor response to these drugs during